The hypothesis posed in our original application was predicated on the concept that tumor cell resistance to DNA-directed chemotherapeutics such as the chloroethylnitrosoureas (CENU), and platinum complexes (cis-Pt and CBDCA) results from DNA repair systems which protect the tumor cell from the cytotoxic DNA lesions. Overwhelming evidence indicated that 0-6 methylguanine DNA methyltransferase (MGMT) was the major contributor to CENU resistance. During the previous funding period we have demonstrated that this DNA repair enzyme can be inhibited by the clinically used agent Streptozotocin (STZ). When highly resistant tumor cells were pretreated with STZ prior to BCNU exposure, virtual complete inhibition of MGMT was obtained, and highly synergistic cell kills were observed. Aim 1 of the current application will continue biochemical modulation of tumor cell resistance to the CENU with MGMT-depleting regimens containing STZ, and the novel MGMT-depleting agent 0-6 Benzylguanine (BG). These studies target the MGMT repair protein in drug resistant cells. Aim 2 will attempt molecular modulation of tumor cell resistance to the CENU using antisense ribozymes transcribed from inducible vectors, to destroy or functionally inhibit long-lived MGMT mRNA. These studies will target long lived MGMT mRNA which may be responsible for reacquisition of CENU resistance in human tumor cells. During the previous funding period we demonstrated that DNA excision repair inhibitors (cytosine arabinoside, Ara-C, and Hydroxyurea, HU) could inhibit the repair of platinum-induced DNA damage, and produce synergistic cell kills. Aim 3 of the current application will attempt biochemical modulation of tumor cell resistance to the platinum compounds using 2',2'Difluorodeoxycytidine (dFdC) and Fludarabine (F-Ara-A) as novel inhibitors of DNA excision repair. These studies will incorporate exciting new compounds in combination with platinum analogs to inhibit platinum damage repair. Aim 4 will characterize the role of ERCC (Excision Repair Cross Complementation) gene expression in tumor cells sensitive and resistant to platinum compounds. The ultimate goal of all of these studies is to increase our understanding of the mechanisms of drug resistance to these agents, and to develop novel regimens to be tested in clinical trials which will attempt to reverse drug resistance in patients.